Deuterated hepatitis C protease inhibitors

ABSTRACT

A deuterated α-ketoamido steric specific compound of the formula  
                 
wherein D denotes a deuterium atom on a steric specific carbon atom.

CROSS-REFERENCE

This application claims the benefits of U.S. Provisional ApplicationSer. No. 60/782,778, filed Mar. 16, 2006, U.S. Provisional ApplicationSer. No. 60/782,976, filed Mar. 16, 2006, and U.S. ProvisionalApplication Ser. No. 60/844,771, filed Sep. 15, 2006.

BACKGROUND OF THE INVENTION

Infection by hepatitis C virus (“HCV”) is a compelling human medicalproblem. HCV is recognized as the causative agent for most cases ofnon-A, non-B hepatitis, with an estimated human sero-prevalence of 3%globally [A. Alberti et al., “Natural History of Hepatitis C,” J.Hepatology, 31., (Suppl. 1), pp. 17-24 (1999)]. Nearly four millionindividuals may be infected in the United States alone [M. J. Alter etal., “The Epidemiology of Viral Hepatitis in the United States,Gastroenterol. Clin. North Am., 23, pp. 437-455 (1994); M. J. Alter“Hepatitis C Virus Infection in the United States,” J. Hepatology, 31.,(Suppl. 1), pp. 88-91 (1999)].

Upon first exposure to HCV only about 20% of infected individualsdevelop acute clinical hepatitis while others appear to resolve theinfection spontaneously. In almost 70% of instances, however, the virusestablishes a chronic infection that persists for decades [S. Iwarson,“The Natural Course of Chronic Hepatitis,” FEMS Microbiology Reviews,14, pp. 201-204 (1994); D. Lavanchy, “Global Surveillance and Control ofHepatitis C,” J. Viral Hepatitis, 6, pp. 35-47 (1999)]. This usuallyresults in recurrent and progressively worsening liver inflammation,which often leads to more severe disease states such as cirrhosis andhepatocellular carcinoma [M. C. Kew, “Hepatitis C and HepatocellularCarcinoma”, FEMS Microbiology Reviews, 14, pp. 211-220 (1994); I. Saitoet. al., “Hepatitis C Virus Infection is Associated with the Developmentof Hepatocellular Carcinoma,” Proc. Natl. Acad. Sci. USA, 87, pp.6547-6549 (1990)]. Unfortunately, there are no broadly effectivetreatments for the debilitating progression of chronic HCV.

The HCV genome encodes a polyprotein of 3010-3033 amino acids [Q. L.Choo, et. al., “Genetic Organization and Diversity of the Hepatitis CVirus.” Proc. Natl. Acad. Sci. USA, 88, pp. 2451-2455 (1991); N. Kato etal., “Molecular Cloning of the Human Hepatitis C Virus Genome FromJapanese Patients with Non-A, Non-B Hepatitis,” Proc. Natl. Acad. Sci.USA, 87, pp. 9524-9528 (1990); A. Takamizawa et. al., “Structure andOrganization of the Hepatitis C Virus Genome Isolated From HumanCarriers,” J. Virol., 65, pp. 1105-1113 (1991)]. The HCV nonstructural(NS) proteins are presumed to provide the essential catalytic machineryfor viral replication. The NS proteins are derived by proteolyticcleavage of the polyprotein [R. Bartenschlager et. al., “NonstructuralProtein 3 of the Hepatitis C Virus Encodes a Serine-Type ProteinaseRequired for Cleavage at the NS3/4 and NS4/5 Junctions,” J. Virol., 67,pp. 3835-3844 (1993); A. Grakoui et. al., “Characterization of theHepatitis C Virus-Encoded Serine Proteinase: Determination ofProteinase-Dependent Polyprotein Cleavage Sites,” J. Virol., 67, pp.2832-2843 (1993); A. Grakoui et. al., “Expression and Identification ofHepatitis C Virus Polyprotein Cleavage Products,” J. Virol., 67, pp.1385-1395 (1993); L. Tomei et. al., “NS3 is a serine protease requiredfor processing of hepatitis C virus polyprotein”, J. Virol., 67, pp.4017-4026 (1993)].

The HCV NS protein 3 (NS3) contains a serine protease activity thathelps process the majority of the viral enzymes, and is thus consideredessential for viral replication and infectivity. It is known thatmutations in the yellow fever virus NS3 protease decrease viralinfectivity [Chambers, T. J. et. al., “Evidence that the N-terminalDomain of Nonstructural Protein NS3 From Yellow Fever Virus is a SerineProtease Responsible for Site-Specific Cleavages in the ViralPolyprotein”, Proc. Natl. Acad. Sci. USA, 87, pp. 8898-8902 (1990)]. Thefirst 181 amino acids of NS3 (residues 1027-1207 of the viralpolyprotein) have been shown to contain the serine protease domain ofNS3 that processes all four downstream sites of the HCV polyprotein [C.Lin et al., “Hepatitis C Virus NS3 Serine Proteinase: Trans-CleavageRequirements and Processing Kinetics”, J. Virol., 68, pp. 8147-8157(1994)].

The HCV NS3 serine protease and its associated cofactor, NS4A, helpprocess all of the viral enzymes, and is thus considered essential forviral replication. This processing appears to be analogous to thatcarried out by the human immunodeficiency virus aspartyl protease, whichis also involved in viral enzyme processing. HIV protease inhibitors,which inhibit viral protein processing, are potent antiviral agents inman indicating that interrupting this stage of the viral life cycleresults in therapeutically active agents. Consequently HCV NS3 serineprotease is also an attractive target for drug discovery.

There are not currently any satisfactory anti-HCV agents or treatments.Until recently, the only established therapy for HCV disease wasinterferon treatment. However, interferons have significant side effects[M. A. Wlaker et al., “Hepatitis C Virus: An Overview of CurrentApproaches and Progress,” DDT, 4, pp. 518-29 (1999); D. Moradpour etal., “Current and Evolving Therapies for Hepatitis C,” Eur. J.Gastroenterol. Hepatol., 11, pp. 1199-1202 (1999); H. L. A. Janssen etal. “Suicide Associated with Alfa-Interferon Therapy for Chronic ViralHepatitis,” J. Hepatol., 21, pp. 241-243 (1994); P. F. Renault et al.,“Side Effects of Alpha Interferon,” Seminars in Liver Disease, 9, pp.273-277. (1989)] and induce long term remission in only a fraction(˜25%) of cases [O. Weiland, “Interferon Therapy in Chronic Hepatitis CVirus Infection”, FEMS Microbiol. Rev., 14, pp. 279-288 (1994)]. Recentintroductions of the pegylated forms of interferon (PEG-INTRON® andPEGASYS®) and the combination therapy of ribavirin and pegylatedinterferon (REBETROL®) have resulted in only modest improvements inremission rates and only partial reductions in side effects. Moreover,the prospects for effective anti-HCV vaccines remain uncertain.

Thus, there is a need for more effective anti-HCV therapies. Suchinhibitors would have therapeutic potential as protease inhibitors,particularly as serine protease inhibitors, and more particularly as HCVNS3 protease inhibitors. Specifically, such compounds may be useful asantiviral agents, particularly as anti-HCV agents.

It was recently discovered that deuterium incorporation in a compoundwill reduce the rate of epimerization via a deuterium isotope effect,thus enhancing the concentration of the active isomers in vivo relativeto its non-deuterated analogs.

SUMMARY OF THE INVENTION

The present invention relates to deuterated compounds of formula (I)

as well as pharmaceutically acceptable salts, prodrugs, and solvatesthereof. In formula (I), D denotes a deuterium atom.

Referring to formula (I),

-   -   D denotes a deuterium atom;    -   R¹ is        in which        is an optionally substituted monocyclic azaheterocyclyl or        optionally substituted multicyclic azaheterocyclyl, or        optionally substituted multicyclic azaheterocyclenyl wherein the        unsaturatation is in the ring distal to the ring bearing the R²¹        moiety and to which the —C(O)—N(R²)—CDR³—C(O)—C(O)—NR⁴R⁵ moiety        is attached;    -   R²¹ is Q³-W³-Q²-W²-Q¹; wherein        -   Each of W² and W³ is independently a bond, —CO—, —CS—,            —C(O)N(Q⁴)-, —CO₂—, —O—, —N(Q⁴)-C(O)—N(Q⁴)-,            —N(Q⁴)-C(S)—N(Q⁴)-, —OC(O)NQ⁴-, —S—, —SO—, —SO₂—, —N(Q⁴)-,            —N(Q⁴)SO₂—, —N(Q⁴)SO₂N(Q⁴)-, and hydrogen when any of W² and            W³ is the terminal group;        -   Each of Q¹, Q², and Q³ is independently a bond, an            optionally substituted aliphatic, an optionally substituted            heteroaliphatic, an optionally substituted cycloaliphatic,            an optionally substituted aryl, an optionally substituted            heteroaryl, an optionally substituted aralkyl, or an            optionally substituted heteroaralkyl; or hydrogen when any            of Q³, Q², or Q¹ is the terminal group, provided that Q² is            not a bond when both W³ and W² are present; and

Each of R², R³, and R⁴, independently, is H or a C₁₋₆ alkyl;

R⁵ is H, alkyl, cycloalkyl, aryl optionally substituted with 1-4 alkylgroups, alkylaryl, aryl, amino optionally substituted with 1 or 2 alkylgroups; and

R²¹ is Q³-W³-Q²-W²-Q¹; wherein each of W² and W³ is independently abond, —CO—, —CS—, —C(O)N(Q⁴)—, —CO₂—, —O—, —N(Q⁴)-C(O)—N(Q⁴)-,—N(Q⁴)-C(S)—N(Q⁴)-, —OC(O)NQ⁴-, —S—, —SO—, —SO₂—, —N(Q⁴)-, —N(Q⁴)SO₂—,—N(Q⁴)SO₂N(Q⁴)-, and hydrogen when any of W² and W³ is the terminalgroup; each of Q¹, Q², and Q³ is independently a bond, an optionallysubstituted aliphatic, an optionally substituted heteroaliphatic, anoptionally substituted cycloaliphatic, an optionally substituted aryl,an optionally substituted heteroaryl, an optionally substituted aralkyl,or an optionally substituted heteroaralkyl; or hydrogen when any of Q³,Q², or Q¹ is the terminal group, provided that Q² is not a bond whenboth W³ and W² are present.

In some embodiments, R¹ is

inwhich

each of R⁶ and R⁸ is independently

-   -   a bond; or    -   optionally substituted (1,1- or 1,2-)cycloalkylene; or    -   optionally substituted (1,1- or 1,2-)heterocyclylene; or    -   methylene or ethylene, substituted with one substituent selected        from the group consisting of an optionally substituted aliphatic        group, an optionally substituted cyclic group and an optionally        substituted aromatic group, and wherein the methylene or        ethylene is further optionally substituted with an aliphatic        group substituent;

each of R⁷, R⁹, and R¹¹ is independently hydrogen or optionallysubstituted aliphatic group;

R¹⁰ is an optionally substituted aliphatic group, optionally substitutedcyclic group or optionally substituted aromatic group;

L is —C(O)—, —OC(O)—, —NR¹¹C(O)—, —S(O)₂—, —NR¹¹S(O)₂—, or a bond; and

n is 0 or 1.

In some embodiments, n is 1.

In some embodiments, R⁶ is methylene substituted with one substituentselected from the group consisting of an optionally substitutedaliphatic group, an optionally substituted cyclic group, and anoptionally substituted aromatic group.

In some embodiments, R⁶ is methylene substituted with isobutyl.

In some embodiments, R⁷ is hydrogen.

In some embodiments, R⁸ is methylene substituted with one substituentselected from the group consisting of an optionally substitutedaliphatic group, an optionally substituted cyclic group, and anoptionally substituted aromatic group. In some other embodiments, R⁸ ismethylene substituted with an optionally substituted cyclic group. Instill some other embodiments, R⁸ is methylene substituted withcyclohexyl.

In some embodiments, R⁹ is hydrogen.

In some embodiments, L is —CO—.

In some embodiments, R¹⁰ is an optionally substituted aromatic group.

In some embodiments, R¹⁰ is selected from the group consisting of

In some embodiments, R¹⁰ is optionally substituted pyrazinyl (e.g.,2-pyrazinyl).

In some embodiments,

is a substituted monocyclic azaheterocyclyl.

In some other embodiments,

is pyrrolidinyl substituted at the 3-position carbon atom withheteroaryloxy, wherein the heteroaryl is further optionally substitutedwith 1-4 halo groups.

In some embodiments,

is

In some embodiments,

is an optionally substituted multicyclic azaheterocyclyl.

In another embodiment,

In some embodiments,

In another embodiment, R² is hydrogen, each of R⁴ and R⁵ independentlyis hydrogen or cyclopropyl. In another embodiment, R³ is propyl. Inanother embodiment, n is 0. In another embodiment, L is —NR¹¹C(O)— andR¹¹ is hydrogen. In another embodiment, R¹⁰ is an optionally substitutedaliphatic group. In another embodiment, R¹⁰ is t-butyl. In anotherembodiment, the compound is

In some embodiments, R¹ is

in which

-   -   A is —(CHX¹)_(a)—;    -   B is —(CHX²)_(b)—;    -   a is 0 to 3;    -   b is 0 to 3, provided that a+b is 2 or 3;    -   each of X¹ and X² is independently selected from hydrogen,        optionally substituted C₁₋₄ aliphatic, and optionally        substituted aryl;    -   each of Y¹ and Y² is independently hydrogen, optionally        substituted aliphatic, optionally substituted aryl, amino, or        —OQ⁴; wherein each Q⁴ is independently hydrogen or an optionally        substituted aliphatic;    -   R²² is an optionally substituted aliphatic, an optionally        substituted heteroaliphatic, an optionally substituted        cycloaliphatic, an optionally substituted heterocycloaliphatic,        an optionally substituted aryl, or an optionally substituted        heteroaryl.In some embodiments, R²¹ is optionally substituted        alkylcarbonyl.

The moiety

includes all of its stereospecific enantiomers, e.g.,

(when A and B are both CH₂, and Y¹ and Y² are both H).

In some embodiments, R²¹ is aminoalkylcarbonyl, haloalkylcarbonyl,arylalkylcarbonyl, arylalkylcarbonyl, cycloaliphaticalkylcarbonyl, orheterocycloaliphaticalkylcarbonyl, each of which is optionallysubstituted with 1-3 substituents. In some embodiments, R²¹ isheterocycloalkyl-oxycarbonylamino-alkylcarbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,bicycloaryl-sulfonylamino-alkylcarbonyl,aryl-alkoxy-carbonylamino-alkyl-carbonyl,alkyl-carbonylamino-alkyl-carbonyl,aliphatic-oxycarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-aminocarbonylamino-alkyl-carbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,alkyl-aminocarbonylamino-alkyl-carbonyl, orbicycloaryl-aminocarbonylamino-alkyl-carbonyl, each of which isoptionally substituted with 1-3 substituents. In some embodiments, R²²is an optionally substituted aliphatic, optionally substitutedheteroaliphatic, optionally substituted cycloaliphatic, optionallysubstituted heterocycloaliphatic, optionally substituted aryl, oroptionally substituted heteroaryl. In some embodiments, R²² isoptionally substituted phenyl, optionally substituted naphthyl,optionally substituted anthracenyl, optionally substituted naphthalene,or optionally substituted anthracene. In some embodiments, each of X¹,X², Y¹ and Y² is hydrogen, each of a and b is 1.

In some embodiments, R²¹ is an optionally substituted alkylcarbonyl.

In some embodiments, R²¹ is an aminoalkylcarbonyl, haloalkylcarbonyl,arylalkylcarbonyl, arylalkylcarbonyl, cycloaliphaticalkylcarbonyl, orheterocycloaliphaticalkylcarbonyl, each of which is optionallysubstituted with 1-3 substituents.

In some embodiments, R²¹ isheterocycloalkyl-oxycarbonylamino-alkylcarbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,bicycloaryl-sulfonylamino-alkylcarbonyl,aryl-alkoxy-carbonylamino-alkyl-carbonyl,alkyl-carbonylamino-alkyl-carbonyl,aliphatic-oxycarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-aminocarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-carbonylamino-alkyl-carbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,alkyl-aminocarbonylamino-alkyl-carbonyl, orbicycloaryl-aminocarbonylamino-alkyl-carbonyl, each of which isoptionally substituted with 1-3 substituents.

In some embodiments, R²² is an optionally substituted aliphatic,optionally substituted heteroaliphatic, optionally substitutedcycloaliphatic, optionally substituted heterocycloaliphatic, optionallysubstituted aryl, or optionally substituted heteroaryl.

In some embodiments, R²² is optionally substituted phenyl, optionallysubstituted naphthyl, optionally substituted anthracenyl, optionallysubstituted naphthalene, or optionally substituted anthracene.

In some embodiments, each of X¹, X², Y¹, and Y² is hydrogen, each of aand b is 1.

In some embodiments, R²² is an optionally substituted aliphatic,optionally substituted heteroaliphatic, optionally substitutedcycloaliphatic, optionally substituted heterocycloaliphatic, optionallysubstituted aryl, or optionally substituted heteroaryl.

In an embodiment, the compound is

The deuterated compounds of this invention undergo slower epimerizationthan its non-deuterated counterparts. As shown below, the deuteratedcompound 1 very slowly converts to a non-deuterated intermediate whichthen converts to epimers 2 and 3. The epimers 2 and 3 then maintain inan equilibrium, which further slows the epimerization of the deuteratedcompound 1.

As a result of their slow epimerization, the deuterated compounds ofthis invention can enhance the concentration of the active isomers invivo relative to its non-deuterated analogs.

In some embodiments, the deuterium enrichment is at least 50% in thecompounds of this invention. In some embodiments, the deuteriumenrichment is at least 80% in the compounds of this invention. In someembodiments, the deuterium enrichment is at least 90% in the compoundsof this invention. In some embodiments, the deuterium enrichment is atleast 99% in the compounds of this invention.

The invention also relates to a pharmaceutical composition comprising apharmaceutically acceptable carrier and a compound of formula (I) or anyof its embodiments described above.

The invention also relates to a method for increasing the concentrationof the active isomer of a pharmaceutical agent in vivo, comprisingadministering to a patient in need thereof a deuterated isomer of thepharmaceutical agent in an amount sufficient to confer thepharmaceutical effect.

The invention also relates to a method for enhancing the bioavailabilityof a compound, comprising replacing a hydrogen atom that is bonded to asteric carbon atom in the compound with a deuterium atom. In oneembodiment, the deuterated compound is of formula (I) or any of itsembodiments described above.

The invention also relates to a method for inhibiting HCV protease,comprising contacting HCV protease with a deuterated compound of formula(I) or any of its embodiments described above.

The invention also relates to a method for treating a patient sufferingfrom HCV infection or a condition mediated by HCV protease, comprisingadministering to the patient a pharmaceutically effective amount of adeuterated compound of formula (I) or any of its embodiments describedabove.

Also within the scope of this invention is a process for preparing anoptically enriched compound of Formula 1, in which

-   -   the carbon atoms alpha and beta to the carboxy group are        stereocenters;    -   R₁ is independently H, optionally substituted aliphatic,        optionally substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic;    -   R′₁ is deuterium,    -   R′₂ is —NHR₂ or —OE;    -   R₂ is H, an optionally substituted aliphatic, optionally        substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic; and    -   E is a C₁₋₆ alkyl or benzyl;

The method includes the steps of:

-   -   a) forming a salt of a compound of Formula 1, and    -   b) crystallizing said salt to give a compound of greater than        55% enantiomeric excess.

In some embodiments, R₁ is C₁₋₆ alkyl, and R′₂ is —NHR₂ wherein R₂ is aC₁₋₆ alkyl or C₁₋₆ cycloalkyl. In some embodiments, R₁ is propyl and R₂is cyclopropyl.

In some embodiments, the method further includes aminating a compound ofFormula ii

with an aminating reagent to provide a compound of Formula iii

In still some embodiments, the aminating reagent is an azide salt andthe intermediate azido compound is reduced by hydrogenation.

In some embodiments, the method further includes oxidizing anunsaturated compound of Formula i

wherein R′₂ is —NHR₂ or —OE, wherein E is C₁₋₅ alkyl or optionallysubstituted benzyl, with an oxidizing reagent to provide a compound ofFormula ii.

In some further embodiments, the oxidizing reagent comprises t-butylhydroperoxide. In some further embodiments, the oxidizing reagentfurther includes a chiral reagent. In some further embodiments, theoxidizing reagent is a mixture of samarium (III) isopropoxide, triphenylarsine oxide, S-(−)1,1′-bi-2-naphthol and 4 Å molecular sieves. In somefurther embodiments, the oxidizing reagent comprises urea-hydrogenperoxide in the presence of trifluoroacetic anhydride.

In some further embodiments, the method further includes hydrolyzing thecompound of Formula ii to give an acid and then converting the acid toan amide compound of Formula ii wherein R′₂ is —NHR₂.

Still within the scope of this invention is a process for preparing acompound of Formula 1

wherein:

-   -   R₁ is H, optionally substituted aliphatic, optionally        substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic;    -   R′₁ is deuterium,    -   R₂ is H, optionally substituted aliphatic, optionally        substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic; and    -   the compound of Formula 1 has an enantiomeric excess of greater        than 55%. The method includes the steps of:    -   a) oxidation of an unsaturated compound of Formula i        to provide a compound of formula ii    -   b) reacting a compound of Formula ii with an aminating reagent        to provide a compound of Formula iii    -   c) forming a salt of a compound of Formula iii with an optically        active organic acid; and    -   d) crystallizing said salt to give a compound of greater than        55% enantiomeric excess.

In some embodiments, the compound of Formula 1 is(2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide. In someembodiments, the organic acid is L-tartaric acid or deoxycholic acid.

Also within the scope of this invention is a process for preparing anoptically enriched compound of Formula 1:

wherein:

-   -   the carbon atoms alpha and beta to the carboxy group are        stereocenters;    -   R₁ is independently H, optionally substituted aliphatic,        optionally substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic;    -   R′₁ is deuterium such that the deuterium enrichment is at least        50%;    -   R′₂ is —NHR₂ or —OE;    -   R₂ is H, optionally substituted aliphatic, optionally        substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic; and    -   E is C₁₋₆ alkyl or benzyl.        The method includes the steps of: a) forming a salt of a        compound of Formula 1, and b) crystallizing said salt to give a        compound of greater than 55% enantiomeric excess.

In some embodiments, R₁ is C₁₋₆ alkyl, and R′₂ is —NHR₂ wherein R₂ is aC₁₋₆ alkyl or C₁₋₆ cycloalkyl. In some embodiments, R₁ is propyl and R₂is cyclopropyl.

In some embodiments, the method further includes the step of aminating acompound of Formula ii

with an aminating reagent to provide a compound of Formula iii

In some embodiments, the aminating reagent is an azide salt and theintermediate azido compound is reduced by hydrogenation.

In some embodiments, the method further includes the step of oxidizingan unsaturated compound of Formula i

wherein R′₂ is —NHR₂ or —OE, wherein E is C₁₋₅ alkyl or optionallysubstituted benzyl, with an oxidizing reagent to provide a compound ofFormula ii.

In some embodiments, the oxidizing reagent comprises t-butylhydroperoxide. In some further embodiments, the oxidizing reagentfurther a chiral reagent. In some embodiments, the oxidizing reagent isa mixture of samarium (III) isopropoxide, triphenyl arsine oxide,S-(−)1,1′-bi-2-naphthol and 4 Å molecular sieves. In some embodiments,the oxidizing reagent comprises urea-hydrogen peroxide in the presenceof trifluoroacetic anhydride.

In some embodiments, R′2 is —OE. In some embodiments, R′₂ is —NHR₂.

In some embodiments, the method further includes hydrolyzing thecompound of Formula ii to give an acid and then converting the acid toan amide compound of Formula ii wherein R′₂ is —NHR₂.

In some embodiments, the method further includes oxidizing a compound ofFormula iv

to give the compound of Formula ii. In some instance, the oxidation isconducted by using manganese dioxide.

In some embodiments, the method further includes reducing a compound ofFormula v

to give the compound of Formula iv. In some instance, the compound isreduced with Red-A1® and then quenched with deuterium oxide. As known inthe art, “Red-A1®” refers to the compound [(CH₃OCH₂OCH₂)₂AlH₂]Na, whichis commercial available, generally as a solution in toluene (e.g., 70%W/W). For more information about Red-A1®, see, e.g., Bates R. W. et al.,Tetrahedron, 1990, 46, 4063.

Still within the scope of this invention is a process for preparing acompound of Formula 1

wherein:

-   -   R₁ is H, optionally substituted aliphatic, optionally        substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic;    -   R′₁ is deuterium,    -   R₂ is H, optionally substituted aliphatic, optionally        substituted cycloaliphatic, optionally substituted        arylaliphatic, optionally substituted heteroaliphatic or        optionally substituted heteroarylaliphatic; and    -   the compound of Formula 1 has an enantiomeric excess of greater        than 55%. The method includes the steps of:    -   a) oxidation of an unsaturated compound of Formula i        to provide a compound of formula ii    -   b) reacting a compound of Formula ii with an aminating reagent        to provide a compound of Formula iii    -   c) forming a salt of a compound of Formula iii with an optically        active organic acid; and    -   d) crystallizing said salt to give a compound of greater than        55% enantiomeric excess.

In some embodiments, the compound of Formula 1 is(2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide. In someembodiments, the organic acid is L-tartaric acid or deoxycholic acid.

DETAILED DESCRIPTION OF THE INVENTION I. DEFINITIONS

A. Terms

As used herein, the term “aliphatic” encompases alkyl, alkenyl andalkynyl.

As used herein, an “alkyl” group refers to a saturated aliphatichydrocarbon group containing 1-8 (e.g., 1-6 or 1-4) carbon atoms. Analkyl group can be straight or branched. Examples of an alkyl groupinclude, but are not limited to, methyl, ethyl, propyl, isopropyl,butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl, and2-ethylhexyl. An alkyl group can be optionally substituted with one ormore substituents such as alkoxy, cycloalkyloxy, heterocycloalkyloxy,aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, amino, nitro,carboxy, cyano, halo, hydroxy, sulfo, mercapto, alkylsulfanyl,alkylsulfinyl, alkylsulfonyl, aminocarbonyl, alkylcarbonylamino,cycloalkylcarbonylamino, cycloalkyl-alkylcarbonylamino,arylcarbonylamino, aralkylcarbonylamino, heterocycloalkyl-carbonylamino,heterocycloalkyl-alkylcarbonylamino, heteroarylcarbonylamino,heteroaralkylcarbonylamino, urea, thiourea, sulfamoyl, sulfamide,alkoxycarbonyl, or alkylcarbonyloxy.

As used herein, an “alkenyl” group refers to an aliphatic carbon groupthat contains 2-8 (e.g., 2-6 or 2-4) carbon atoms and at least onedouble bond. Like an alkyl group, an alkenyl group can be straight orbranched. Examples of an alkenyl group include, but are not limited to,allyl, isoprenyl, 2-butenyl, and 2-hexenyl. An alkenyl group can beoptionally substituted with one or more substituents such as alkoxy,cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,heteroarylalkoxy, amino, nitro, carboxy, cyano, halo, hydroxy, sulfo,mercapto, alkylsulfanyl, alkylsulfinyl, alkylsulfonyl, aminocarbonyl,alkylcarbonylamino, cycloalkylcarbonylamino,cycloalkyl-alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino,heterocycloalkyl-carbonylamino, heterocycloalkyl-alkylcarbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, urea, thiourea,sulfamoyl, sulfamide, alkoxycarbonyl, or alkylcarbonyloxy.

As used herein, an “alkynyl” group refers to an aliphatic carbon groupthat contains 2-8 (e.g., 2-6 or 2-4) carbon atoms and has at least onetriple bond. An alkynyl group can be straight or branched. Examples ofan alkynyl group include, but are not limited to, propargyl and butynyl.An alkynyl group can be optionally substituted with one or moresubstituents such as alkoxy, cycloalkyloxy, heterocycloalkyloxy,aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, amino, nitro,carboxy, cyano, halo, hydroxy, sulfo, mercapto, alkylsulfanyl,alkylsulfinyl, alkylsulfonyl, aminocarbonyl, alkylcarbonylamino,cycloalkylcarbonylamino, cycloalkyl-alkylcarbonylamino,arylcarbonylamino, aralkylcarbonylamino, heterocycloalkyl-carbonylamino,heterocycloalkyl-alkylcarbonylamino, heteroarylcarbonylamino,heteroaralkylcarbonylamino, urea, thiourea, sulfamoyl, sulfamide,alkoxycarbonyl, or alkylcarbonyloxy.

As used herein, an “amino” group refers to —NR^(X)R^(Y) wherein each ofR^(X) and R^(Y) is independently hydrogen, alkyl, cycloalkyl,(cycloalkyl)alkyl, aryl, aralkyl, heterocycloalkyl,(heterocycloalkyl)alkyl, heteroaryl, or heteroaralkyl. When the term“amino” is not the terminal group (e.g., alkylcarbonylamino), it isrepresented by —NR^(X)—. R^(X) has the same meaning as defined above.

As used herein, an “aryl” group refers to phenyl, naphthyl, or abenzofused group having 2 to 3 rings. For example, a benzofused groupincludes phenyl fused with one or two C₄₋₈ cycloaliphatic moieties,e.g., 1, 2, 3, 4-tetrahydronaphthyl, indanyl, dihydroindanyl, orfluorenyl. An aryl is optionally substituted with one or moresubstituents such as alkyl (including carboxyalkyl, hydroxyalkyl, andhaloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl,(cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl,heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy,heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, amino,nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl,alkylcarbonylamino, cycloalkylcarbonylamino,(cycloalkyl)alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkyl)alkylcarbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

As used herein, an “aralkyl” group refers to an alkyl group (e.g., aC₁₋₄ alkyl group) that is substituted with an aryl group. Both “alkyl”and “aryl” have been defined above. An example of an aralkyl group isbenzyl.

As used herein, cycloaliphatic encompasses cycloalkyl, cycloalkenyl andcycloalkynyl.

As used herein, a “cycloalkyl” group refers to an aliphatic carbocyclicring of 3-10 (e.g., 4-8) carbon atoms. Examples of cycloalkyl groupsinclude cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl,norbornyl, cubyl, octahydro-indenyl, decahydro-naphthyl,bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl, andbicyclo[3.2.3]nonyl,. A “cycloalkenyl” group, as used herein, refers toa non-aromatic carbocyclic ring of 3-10 (e.g., 4-8) carbon atoms havingone or more double bonds. Examples of cycloalkenyl groups includecyclopentenyl, 1,4-cyclohexa-di-enyl, cycloheptenyl, cyclooctenyl,hexahydro-indenyl, octahydro-naphthyl, bicyclo[2.2.2]octenyl, andbicyclo[3.3.1]nonenyl. A cycloalkyl or cycloalkenyl group can beoptionally substituted with one or more substituents such as alkyl(including carboxyalkyl, hydroxyalkyl, and haloalkyl such astrifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl,heterocycloalkyl, (heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy,cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro, carboxy,alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino,cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino,arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkyl)alkylcarbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

As used herein, heterocycloaliphatic means heterocycloalkyl,heterocycloalkenyl and heterocycloalkynyl. As used herein, a“heterocycloalkyl” group refers to a 3- to 10-membered (e.g., 4- to8-membered) saturated ring structure, in which one or more of the ringatoms is a heteroatom, e.g., N, O, or S. Examples of a heterocycloalkylgroup include piperidinyl, piperazinyl, tetrahydropyranyl,tetrahydrofuryl, dioxolanyl, oxazolidinyl, isooxazolidinyl, morpholinyl,octahydro-benzofuryl, octahydro-chromenyl, octahydro-thiochromenyl,octahydro-indolyl, octahydro-pyrindinyl, decahydro-quinolinyl,octahydro-benzo[b]thiophenyl, 2-oxa-bicyclo[2.2.2]octyl,1-aza-bicyclo[2.2.2]octyl, 3-aza-bicyclo[3.2.1]octyl, anad2,6-dioxa-tricyclo[3.3.1.03,7]nonyl. A “heterocycloalkenyl” group, asused herein, refers to a 3- to 10-membered (e.g., 4- to 8-membered)non-aromatic ring structure having one or more double bonds, and whereinone or more of the ring atoms is a heteroatom, e.g., N, O, or S. Aheterocycloalkyl or heterocycloalkenyl group can be optionallysubstituted with one or more substituents such as alkyl (includingcarboxyalkyl, hydroxyalkyl, and haloalkyl such as trifluoromethyl),alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl, heterocycloalkyl,(heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy,heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro, carboxy,alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino,cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino,arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkyl)alkylcarbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl. In some instances, asubstituent on the heterocycloalkyl or heterocycloalkenyl itself can becyclic (which optionally contains one or more hetero atoms) such thatthe resultant substituted heterocycloalkyl orheterocycloalkenyl is a spiro ring system, e.g.,

A “heteroaryl” group, as used herein, refers to a monocyclic, bicyclic,or tricyclic ring structure having 5 to 15 ring atoms wherein one ormore of the ring atoms is a heteroatom, e.g., N, O, or S and wherein oneor more rings of the bicyclic or tricyclic ring structure is aromatic.Some examples of heteroaryl are pyridyl, furyl, pyrrolyl, thienyl,thiazolyl, oxazolyl, imidazolyl, indolyl, 2,3-dihydroindolyl, quinolyl,1,2-dihydroquinolyl, 1,2,3,4-tetrahydroquinolyl, tetrazolyl, benzofuryl,2,3-dihydrobenzofuranyl, benzthiazolyl, xanthene, thioxanthene,phenothiazine, dihydroindole, and benzo[1,3]dioxole. A heteroaryl isoptionally substituted with one or more substituents such as alkyl(including carboxyalkyl, hydroxyalkyl, and haloalkyl such astrifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl,heterocycloalkyl, (heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy,cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro, carboxy,alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino,cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino,arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkyl)alkylcarbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

A “heteroaralkyl” group, as used herein, refers to an alkyl group (e.g.,a C₁₋₄ alkyl group) that is substituted with a heteroaryl group. Both“alkyl” and “heteroaryl” have been defined above.

As used herein, “cyclic moiety” includes cycloalkyl, heterocycloalkyl,cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl, each of which hasbeen defined previously.

As used herein, an “acyl” group refers to a formyl group or alkyl-C(═O)—where “alkyl” has been defined previously. Acetyl and pivaloyl areexamples of acyl groups.

As used herein, a “carbamoyl” group refers to a group having thestructure —O—CO—NR^(x)R^(y) or —NR^(x)—CO—O—R^(z) wherein R^(x) andR^(y) have been defined above and R^(z) can be alkyl, aryl, aralkyl,heterocycloalkyl, heteroaryl, or heteroaralkyl.

As used herein, a “carboxy” and a “sulfo” group refer to —COOH and—SO₃H, respectively.

As used herein, an “alkoxy” group refers to an alkyl-O— group where“alkyl” has been defined previously.

As used herein, a “sulfoxy” group refers to —O—SO—R^(X) or —SO—O—R^(X),where R^(X) has been defined above.

As used herein, a sulfanyl group refers to —S—R^(X), where R^(X) hasbeen defined above.

As used herein, a sulfinyl group refers to —S(O)—R^(X), where R^(X) hasbeen defined above.

As used herein, a sulfonyl group refers to —S(O)₂—R^(X), where R^(X) hasbeen defined above.

As used herein, a “halogen” or “halo” group refers to fluorine,chlorine, bromine or iodine.

As used herein, a “sulfamoyl” group refers to the structure—S(O)₂—NR^(x)R^(y) or —NR^(x)—S(O)₂—R^(z) wherein R^(x), R^(y), andR^(z) have been defined above.

As used herein, a “sulfamide” group refers to the structure—NR^(X)—S(O)₂—NR^(Y)R^(Z) wherein R^(X), R^(Y), and R^(Z) have beendefined above.

As used herein, a “urea” group refers to the structure—NR^(X)—CO—NR^(Y)R^(Z) and a “thiourea” group refers to the structure—NR^(X)—CS—NR^(Y)R^(Z). R^(X), R^(Y), and R^(Z) have been defined above.

As used herein, a “guanidino” group refers to the structure—N═C(NR^(x)R^(y))N(R^(x)R^(y)) wherein R^(X) and R^(Z) have been definedabove.

As used herein, the term “amidino group” refers to the structure—C═(NR^(x))N(R^(x)R^(y)) wherein R^(X) and R^(Y) have been definedabove.

As used herein, the term “oximino group” refers to the structure—C═N—OR^(x) wherein R^(x) has been defined above.

As used herein, an effective amount is defined as the amount required toconfer a therapeutic effect on the treated patient, and is typicallydetermined based on age, surface area, weight, and condition of thepatient. The interrelationship of dosages for animals and humans (basedon milligrams per meter squared of body surface) is described byFreireich et al., Cancer Chemother. Rep., 50: 219 (1966). Body surfacearea may be approximately determined from height and weight of thepatient. See, e.g., Scientific Tables, Geigy Pharmaceuticals, Ardsley,N.Y., 537 (1970).

As used herein, “patient” refers to a mammal, including a human.

An antagonist, as used herein, is a molecule that binds to the receptorwithout activating the receptor. It competes with the endogenousligand(s) or substrate(s) for binding site(s) on the receptor and, thusinhibits the ability of the receptor to transduce an intracellularsignal in response to endogenous ligand binding.

The phrase “optionally substituted” is used interchangeably with thephrase “substituted or unsubstituted.” As described herein, compounds ofthe invention may optionally be substituted with one or moresubstituents, such as are illustrated generally above, or as exemplifiedby particular classes, subclasses, and species of the invention. Unlessotherwise noted, each of the specific groups for the variables R¹, R²,R³, R⁴, and R⁵ in formula (I) may be optionally substituted with one ormore substituents described herein. Each substituent of a specific groupis further optionally substituted with one to three of halo, cyano,alkoxy, hydroxyl, nitro, haloalkyl, and alkyl. For instance, an alkylgroup may be substituted with alkylsulfanyl and the alkylsulfanyl may beoptionally substituted with one to three of halo, oxo, cyano, alkoxy,hydroxyl, nitro, haloalkyl, and alkyl. As an additional example, analkyl may be substituted with a (cycloalkyl)carbonylamino and thecycloalkyl portion of a (cycloalkyl)carbonylamino may be optionallysubstituted with one to three of halo, cyano, oxo, alkoxy, hydroxyl,nitro, haloalkyl, and alkyl.

In general, the term “substituted,” whether preceded by the term“optionally” or not, refers to the replacement of hydrogen radicals in agiven structure with the radical of a specified substituent. Specificsubstituents are described above in the definitions and below in thedescription of compounds and examples thereof. Unless otherwiseindicated, an optionally substituted group may have a substituent ateach substitutable position of the group, and when more than oneposition in any given structure may be substituted with more than onesubstituent selected from a specified group, the substituent may beeither the same or different at every position. A ring substituent, suchas a heterocycloalkyl, may be bound to another ring, such as acycloalkyl, to form a spiro-bicyclic ring system, e.g., both rings shareone common atom. As one of ordinary skill in the art will recognize,combinations of substituents envisioned by this invention are thosecombinations that result in the formation of stable or chemicallyfeasible compounds.

The phrase “stable or chemically feasible,” as used herein, refers tocompounds that are not substantially altered when subjected toconditions to allow for their production, detection, and preferablytheir recovery, purification, and use for one or more of the purposesdisclosed herein. In some embodiments, a stable compound or chemicallyfeasible compound is one that is not substantially altered when kept ata temperature of 40° C. or less, in the absence of moisture or otherchemically reactive conditions, for at least a week.

Unless otherwise stated, structures depicted herein are also meant toinclude all isomeric (e.g., enantiomeric, diastereomeric, and geometric(or conformational)) forms of the structure; for example, the R and Sconfigurations for each asymmetric center, (Z) and (E) double bondisomers, and (Z) and (E) conformational isomers. Therefore, singlestereochemical isomers as well as enantiomeric, diastereomeric, andgeometric (or conformational) mixtures of the present compounds arewithin the scope of the invention. Unless otherwise stated, alltautomeric forms of the compounds of the invention are within the scopeof the invention.

Additionally, unless otherwise stated, structures depicted herein arealso meant to include compounds that differ only in the presence of oneor more isotopically enriched atoms. For example, compounds having thepresent structures except for the replacement of hydrogen by deuteriumor tritium, or the replacement of a carbon by a ¹³C- or ¹⁴C-enrichedcarbon are within the scope of this invention. Such compounds areuseful, for example, as analytical tools or probes in biological assays.

An N-oxide derivative or a pharmaceutically acceptable salt of each ofthe compounds of formula (I) is also within the scope of this invention.For example, a nitrogen ring atom of the imidazole or pyrazole core ringor a nitrogen-containing heterocyclyl substituent can form an oxide inthe presence of a suitable oxidizing agent such as m-chloroperbenzoicacid or H₂O₂.

A compound of formula (I) that is acidic in nature (e.g., having acarboxyl or phenolic hydroxyl group) can form a pharmaceuticallyacceptable salt such as a sodium, potassium, calcium, or gold salt. Alsowithin the scope of the invention are salts formed with pharmaceuticallyacceptable amines such as ammonia, alkyl amines, hydroxyalkylamines, andN-methylglycamine. A compound of formula (I) can be treated with an acidto form acid addition salts. Examples of such acids include hydrochloricacid, hydrobromic acid, hydroiodic acid, sulfuric acid, methanesulfonicacid, phosphoric acid, p-bromophenyl-sulfonic acid, carbonic acid,succinic acid, citric acid, benzoic acid, oxalic acid, malonic acid,salicylic acid, malic acid, fumaric acid, ascorbic acid, maleic acid,acetic acid, and other mineral and organic acids well known to thoseskilled in the art. The acid addition salts can be prepared by treatinga compound of formula (I) in its free base form with a sufficient amountof an acid (e.g., hydrochloric acid) to produce an acid addition salt(e.g., a hydrochloride salt). The acid addition salt can be convertedback to its free base form by treating the salt with a suitable diluteaqueous basic solution (e.g., sodium hydroxide, sodium bicarbonate,potassium carbonate, or ammonia). Compounds of formula (I) can also be,for example, in a form of achiral compounds, racemic mixtures, opticallyactive compounds, pure diastereomers, or a mixture of diastereomers.

B. Abbreviations

The following abbreviations have the following meanings. If anabbreviation is not defined, it has its generally accepted meaning.

BEMP=2-tert-butylimino-2-diethylamino-1,3-dimethylperhydro-1,3,2-diazaphosphorine

Boc=t-butoxycarbonyl

BOP=benzotriazol-1-yloxy-tris(dimethylamino)phosphoniumhexafluorophosphate

bd=broad doublet

bs=broad singlet

CDI=carbonyl diimidazole

d=doublet

dd=doublet of doublets

DIC=diisopropylcarbodiimide

DMF=dimethylformamide

DMAP=dimethylaminopyridine

DMSO=dimethylsulfoxide

EDCI=ethyl-1-(3-dimethyaminopropyl)carbodiimide

eq.=equivalents

EtOAc=ethyl acetate

g=grams

HOBT=1-hydroxybenzotriazole

DIPEA=Hunig's base=diisopropylethylamine

L=liter

m=multiplet

M=molar

max=maximum

meq=milliequivalent

mg=milligram

mL=milliliter

mm=millimeter

mmol=millimole

MOC=methoxyoxycarbonyl

N=normal

N/A=not available

ng=nanogram

nm=nanometers

OD=optical density

PEPC=1-(3-(1-pyrrolidinyl)propyl)-3-ethylcarbodiimide

PP-HOBT=piperidine-piperidine-1-hydroxybenzotrizole

psi=pounds per square inch

Ph=phenyl

q=quartet

quint.=quintet

rpm=rotations per minute

s=singlet

t=triplet

TFA=trifluoroacetic acid

THF=tetrahydrofuran

tlc=thin layer chromatography

μL=microliter

UV=ultra-violet

II. COMPOUNDS OF THIS INVENTION

Generally, the deuterated compounds of this invention can be synthesizedby methods known in the art as for synthesizing their non-deuteratedforms, except that a deuterated starting material or a reacting reagentis used during the synthesis process. Examples of applicable methodsinclude those described in U.S. Application No. 60/711,530; WO 02/18369;WO 07/022459; Advanced Organic Chemistry, 2^(nd) Ed., p. 204, J. March,McGraw Hill, New York, N.Y., 1997; and Synthesis of A: Elemes andRagnarsosson, J. of Chem. Soc., Perkin 1, 1996, 537.

All publications cited herein are incorporated by reference in theirentireties.

Compounds of Formula I are prepared using known methods, for example,such as illustrated below in Scheme I.

Referring to Scheme I, the acid of Formula i is reacted with adeuterated amino-alcohol-amide of Formula ii in the presence of acondensing reagent such as, for example, EDCI and HOSu to provide thehydroxy-amide of Formula iii. In some embodiments, the percent deuterium(D) enrichment as shown in ii is greater than 10%. In other embodimentsthe enrichment is from 10% to 99.95%, 40% to 99.95%, 50% to 99.95%, 60%to 99.95%, 80% to 99.95%, 90% to 99.95%, 93% to 99.95%, 97% to 99.95%,or 99-99.95%, or 99.95% or higher. Oxidation of iii with a suitableoxidizing reagent provides the compounds of Formula I. Suitableoxidizing reagents include, for example, Dess-Martin periodinane orTEMPO and sodium hypochlorite.

The deuterated amino-alcohol-amides of Formula ii shown in Scheme 1 canbe prepared by using known methods and, for example, as illustratedbelow in Scheme II.

Referring to Scheme II, conversion of the glycine iminic ester iv to thedeuterated sultam of Formula vii is conducted according to procedurespreviously described (Y. Elemes and U. Ragnarsson, J. Chem. Soc., PerkinI, 1996, 6, p. 537. Consecutive treatment of compounds of Formula viiwith acid and base as previously described (L. Lankiewicz, et. al., J.Chem. Soc., Perkin I, 1994, 17, p. 2503 followed by treatment of theintermediate amino acid (not shown) with benzyloxycarbonyl chlorideprovides the protected deuterated amino acid viii. Reaction of viii withmethoxymethylamine in the presence of the condensing reagent CDIprovides the Weinreb amide of Formula ix. Reduction of vi with, forexample, diisobutylaluminum hydride or lithium aluminum hydride providesthe aldehyde x. Using procedures similar to those previously described(see, e.g., WO 02/18369), the aldehyde x is converted to the cyanohydrinxi and thence to the protected hydroxy-amino acid xii. The acid xii isconverted to the protected amide xiii which is deprotected to providethe amino-amide ii.

Alternatively, the deuterated amino-amide ii depicted in Scheme I,wherein R₂ is H, may be prepared, e.g., as illustrated in Scheme III.

Referring to Scheme III, the propargyl alcohol xiv is reduced withsodium bis(2-methoxyethoxy)aluminumhydride, followed by qenching thereaction mixture with deuterium oxide to provide the deuterated allylicalcohol xv. Oxidation of xv with manganese dioxide provides the aldehydexvi which is further oxidized to the acid xvii with sodium chlorite(NaClO₂) in the presence of sodium phosphate and 2-methyl-2-butene.Reaction of the acid xvii with isobutylchloroformate (ICBF) in thepresence of N-methylmorpholine followed by reaction of the intermediatemixed anhydride with the amine HNR₄R₅ provides the amide xviii.Epoxidation of xviii to provide the epoxide xix may be acheived withurea hydrogen peroxide (UHP) in the presence of trifluoracetic acid andp-toluenesulfonic acid. Reaction of xix with sodium azide provides theintermediate azido compound xx which is subsequently reduced to theracemic-aminoalcohol xxi by catalytic hydrogenation in the presence ofpalladium on carbon. The racemic aminoalcohol xxi may be resolved usingknown methods such as chiral chromatography, preparation of opticallyactive derivatives or the formation of salts with an optically activeacid HA followed by crystallization from an organic solvent. Suitableoptically active organic acids for preparing salts include, for example,L-tartaric acid, L-malic acid, (S)-mandelic acid,(1S)-(+)-10-camphorsulfonic acid,(−)2,2:4,6-di-O-isopropylidiene-2-keto-L-gulonic acid hydrate,N-acety-L-leucine, deoxycholic acid, (+)-O,O′-dibenzoyl-D-tartaric acid,O,O′-di-(4-toluoyl)-D-tartaric acid, S-(+)1,1-binaphtyl-2-2-phosphoricacid, L-lactic acid, D-Gluconic acid, lactobionic acid,dipivaloyl-L-tartaric acid, S-(+)-O-acetylmandelic acid andS-(−)2-(phenylcarbamoyloxy)propionic acid. Examples of suitable organicsolvents for recrystallization include dimethylacetamide, ethyl acetateand acetone.

The deuterated compounds thus obtained can be characterized byconventional analytical methods, e.g., NMR and Mass Spectroscope. NMRcan be used to determine a compound's structure, while Mass Spectroscopycan be used to determine the amount of deuterium atom in the compound bycomparison with its non-deuterated form.

The deuterated compounds of this invention are generally more stable andless inclined to epimerize than their non-deuterated analogs. Thus, theycan be used in application where specific steric configuration in thecompounds of this invention is desired. For instance, the deuteratedcompounds of formula (I) may be used to treat or prevent infectioncaused by HCV or other HCV protease-mediated condition, as they arecapable of inhibiting HCV protease. Their inhibitory activity can bemeasured by traditional enzyme inhibition essays, some of which aredescribed in the publications cited above. See, e.g., Perni, R. B. etal., Antimicrobial Agents and Chemotherapy, 2006 (march), 50 (3):899-909.

Additionally, the deuterated compounds of formula (I) can be used as abiological tool to study the pharmacological properties of theirnon-deuterated analogs. Accordingly, these uses are also within thescope of this invention.

EXAMPLE 1 Preparation of(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)-N—((S)-1-(cyclopropylamino)-1,2-dioxo-3-deutero-hexan-3-yl)octahydrocyclopenta[c]pyrrole-1-carboxamide

Step a: Preparation of

The deuterated sultam (i.e., compound vi shown in the scheme below) wasprepared by known methods such as those described in Y. Elemes and U.Ragnarsson, J. of Chem. Soc., Perkin 1, 1996, 6, 537; W. Oppolzer,et.al., Helv. Chim. Acta., 1994, 25: 2363, by using the correspondingunsubstituted sultam and propyl iodide.

17.32 g of compound vi (45.8 mmol) and 229 mL of THF were then chargedinto a 500 mL round-bottomed-flask with a magnetic stir bar and N₂inlet. The resulting solution was cooled to −78° C. and n-BuLi (31.5 mLof a 1.6 M solution in hexane, 50.3 mmol) was added with a syringe pumpover an hour. The resulting yellow solution was aged for 30 minutesbefore a solution of HPMA (56 mL) and n-PrI (13.4 mL, 137 mmol) wasadded to it over 30 minutes. The mixture was allowed to warm to the roomtemperature over 8 hours and then cooled to −20° C. before D₂O (50 mL)was added to the mixture. The reaction mixture was then extracted withEtOAc (400 mL) and the organic layer was dried over MgSO4 andconcentrated to provide 61.3 g of the crude oil. Chromatography on 500 gof silica gel eluting with 2:1 heptane/EtOAc followed by concentrationof the rich cut gave 20.35 g of a white solid. The white solid was thenrecrystallized from EtOH (210 mL) to give 15.39 g of compound vii as awhite crystalline solid. The deuterium incorporation was 93% asdetermined by ¹H NMR.

Step b: Preparation of (S)-2-(benzyloxycarbonylamino)-2-deuteropentanoicacid, viii

Compound vii (15.39 g, 32.1 mmol) from step a was charged into THF (100mL) and 1N HCl (50 mL). The resulting emulsion was stirred overnight atthe room temperature and then concentrated under vacuum to provide athick oil. The oil was then dissolved in THF (100 mL), and to thesolution was added water (25 mL) and LiOH (3.08 g, 128 mmol). Thissolution was stirred overnight again at the room temperature and thenconcentrated to remove THF. A hazy light yellow emulsion remained. Thiswas diluted with water (25 mL) and extracted with CH₂Cl₂ (three times,50 mL each). The aqueous phase was diluted with THF (200 mL) and cooledto 0° C. while stirring rapidly and CBZ-Cl (7.6 mL, 54 mmol) was addeddropwise over 15 minutes. After stirring for an hour at 0° C., the THFsolvent was removed in vacuo and the residue was acidified by additionof 1N HCl (50 mL). The solution was extracted with EtOAc (3 times, 100mL each) and the organic phase was dried over Na₂SO₄ and concentrated toprovide an oil. The residue was dissolved in EtOAc (25 mL) and heptane(150 mL), seeded and stirred overnight at the room temperature. Thesolids were collected on a flit, rinsed with heptane (30 mL) and airdried to give 5.65 g (70%) of compound viii shown in the scheme above.The deuterium incorporation was 93% as determined by ¹H NMR.

Step c: Preparation of (S)-benzyl1-(methoxy(methyl)amino)-1-oxo-2-deuteropentan-2-ylcarbamate

To a flask containing 1.0 g of(S)-2-(benzyloxycarbonylamino)-2-deuteropentanoic acid (3.97 mmol) in 20mL of dichloromethane maintained at 0° C., was added 3.0 eq. ofN-methylmorpholine (700 uL), 1.5 eq. of N,O-dimethylhydroxylaminehydrochloride (581 mg) and 1.5 eq. of EDCI (1.14 g). The reactionmixture was stirred overnight from 0° C. to the room temperature. Thereaction mixture was then diluted in dichloromethane and washed with HCl(1N) and brine. The organic layer was dried over MgSO₄. The crudemixture was purified by flash chromatography (ethyl acetate 15-75% inhexanes) to afford 814 mg of pure amide (title compound). ES+=296.1,ES−=295.2. ¹H NMR spectrum confirmed the structure.

Step d: Preparation of (S)-benzyl 1-oxo-2-deuteropentan-2-ylcarbamate

Using procedures described in WO 02/18369, the Cbz-protected amino acidof Step c is converted to the title compound. Specifically, into a flaskcontaining 1.0 eq. of (S)-benzyl1-(methoxy(methyl)amino)-1-oxo-2-deuteropentan-2-ylcarbamate (810 mg,2.75 mmol) in 10 mL of dry THF maintained at 0° C. (in an ice bath) wasadded slowly 1.7 eq. of a solution of lithium borohydride (1.0M) (4.67mL). After about 10 minutes, the ice bath was removed and the reactioncontinue for an hour. The reaction was quenched at 0° C. by adding 5 mLof a solution of KHSO₄ (10%). The solution was then diluted by theaddition of 10 mL of HCl (1N). The mixture was stirred for 30 minutes,then extracted 3 times with dichloromethane. The organic phases werecombined and washed with a solution of HCl (1 N), water and brine. Theorganic phase was then dried over MgSO₄ and the volatile evaporated. Thealdehyde was used as is in the next step. ES+=237.1, ES−=235.2.

Step e: Preparation of benzyl(3S)-1-(cyclopropylamino)-2-hydroxy-1-oxo-3-deuterohexan-3-ylcarbamate

Cyclopropyl isonitrile was prepared according to the scheme shown below.

The cyclopropyl isonitrile was then coupled with the aldehyde product ofStep d to give the title compound as described in J. E. Semple et al.,Org. Lett., 2000, 2(18), p. 2769; Lumma W., J. Org. Chem., 1981, 46,3668″. ES+=322.1.

Step f: Preparation of(3S)-3-amino-N-cyclopropyl-3-deutero-2-hydroxyhexanamide

Hydrogenolysis of the Cbz compound of Step e was achieved by using apalladium on carbon catalyst in the presence of hydrogen to give thetitle compound. Shown in the following schemes are Steps c, d, e, and f.

Step g: Preparation of(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)-N-((3S)-1-(cyclopropylamino)-3-deutero-2-hydroxy-1-oxohexan-3-yl)octahydrocyclopenta[c]pyrrole-1-carboxamide

The title compound was prepared from the hydroxy-amino amide product ofStep f by condensation with the appropriate acid in the presence of acoupling reagent such as, e.g., EDCI and HOSu. Specifically, in a flaskcontaining 1.2 eq. of(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)octahydrocyclopenta[c]pyrrole-1-carboxylicacid (1.59 g) in 20 mL of DMF, was added 2.5 eq. of diisopropylamine(980 uL), 1.2 eq. N-hydroxybenzotriazole hydrate (411 mg) and 1.3 eq. ofEDCI (558 mg). After 15 minutes of stirring at the room temperature, 1.0eq. of (3S)-3-amino-N-cyclopropyl-3-deutero-2-hydroxyhexanamidehydrochloride (500 mg) was added to the mixture. After another 24 hours,the reaction mixture was diluted into 400 mL of ethyl acetate. Theorganic phase of the mixture was washed with HCl (1N), water, saturatedsodium bicarbonate solution, brine, and then dried over MgSO₄. The crudeproduct was purified by chromatography on silica (ethyl acetate 70-100%in Hexanes) to give 1.31 g of the tile compound as a white solid.ES+=683.6, ES−=682.2. The NMR ¹H confirmed the structure.

Step h: Preparation of(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)-N—((S)-1-(cyclopropylamino)-1,2-dioxo-3-deutero-hexan-3-yl)octahydrocyclopenta[c]pyrrole-1-carboxamide

The title compound was prepared by oxidation of the product of Step gwith a suitable oxidizing reagent such as Dess Martin periodinane orTEMPO and sodium hypochlorite. Specifically, in a flask containing 1.31g of(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)-N-((3S)-1-(cyclopropylamino)-3-deutero-2-hydroxy-1-oxohexan-3-yl)octahydrocyclopenta[c]pyrrole-1-carboxamidein 40 mL of dichloromethane was added at room temperature 1.06 g of DessMartin periodinane. After 2 hours of stirring, 50 mL of sodium bisulfite(1N) was added, and the mixture was stirred for 30 minutes. The 2 phaseswere separated, the organic was washed with water twice, brine and driedover Na2SO4. The crude product was purified by chromatography on silica(ethyl acetate 20-100% in Hexanes) to give 1.07 g of the title compoundas a white solid. ES+=681.5, ES−=680.0. The ¹H NMR spectrum confirmedthe structure.

Deuterium incorporation was determined by MS to be 93%. Thediastereoisome ratio was determined by chiral HPLC normal phase and washigher than 99% d.e.

The following scheme shows the reactions of both Steps g and h.

EXAMPLE 2 Preparation of(2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamidehydrochloride

The scheme shown above illustrate the total synthesis of the titlecompound. Each step is described in detail as follows.

Step 1: Preparation of 3-deutero-(E)-hex-2-en-1-ol

To a three-neck 250 mL round bottom flask equipped with mechanicalstirrer and reflux condenser was charged 2-hexyn-1-ol (10 g, 0.1 mole)and THF (100 mL, 10 vol). The resulting mixture was cooled to 0±5° C.and then Red-A1 (65% in Toluene, 32 mL, 1.6 eq) was added slowly under anitrogen atmosphere between 0° C. and 20° C. The resulting mixture wasallowed to be warmed up to 25° C. and stirred for 5 hours. The reactionmixture was cooled down to −5±5° C. and D₂O (8.2 g, 4 eq.) was addeddrop wise between 0° C. and 15° C. To the resulting mixture was chargedIPAC (50 mL, 5 vol) and saturated NH₄Cl solution (50 mL, 5 vol.). Afterstirring the mixture for 10 min, the white solid formed was filteredout. The organic layer from the filtrate was separated and the aqueouslayer was extracted with IPAC(30 mL, 3 vol). The organic layers werecombined and washed with water (30 mL, 3 vol) and dried over MgSO₄ andconcentrated to afford 9.8 g of the product (compound 2) as a colorlessoil. The crude product 2 was used for the next step without furtherpurification.

¹H NMR (500 MHz, CDCl₃) δ 5.66 (t, 1H, J=5.0 Hz), 4.12 (d, 2H, J=5.0Hz), 2.04 (t, 2H, J=5.0 Hz), 1.38˜1.46 (m, 2H), 0.93 (t, 3H, J=5.0 Hz)

Step2: Preparation of 3-deutero-(E)-hex-2-enal

To a three-neck 250 mL round bottom flask equipped with mechanicalstirrer containing 3-deutero-2-hexenol (10 g, 0.1 mole) in CH₂Cl₂ (150mL, 15 vol) was charged activated MnO₂ (87 g, 10 eq) at roomtemperature. After vigorous stirring for 1 hour, another portion of MnO₂(16 g, 2 eq) was added and the shaking was continued for 4 hours. Thereaction solution was filtered through a pad of celite. The solvent wasremoved in vacuo (25° C., 100 mmHg) to give 8.8 g of the crude aldehydeproduct (compound 3) as a pale yellowish oil. The crude product was usedfor the next step without further purification.

¹H NMR (500 MHz, CDCl₃) δ 9.54 (d, 1H, J=10.0 Hz), 6.14(s, 1H), 2.34 (m,2H), 1.55˜1.60 (m, 2H), 1.00 (t, 3H, J=5.0 Hz)

Step 3: Preparation of 3-deutero-(E)-hex-2-enoic acid

To a three-neck 500 mL round bottom flask equipped with mechanicalstirrer and reflux condenser was charged 3-deutero-2-Hexen-1-al (10 g,0.1 mole), tert-BuOH (90 mL, 9 vol), and 2-methyl-2-butene (30 mL, 3vol). The resulting solution was added with a freshly prepared aqueousNaClO₂ (27.4 g, 3 eq) and NaH₂PO₄ (62.9 g, 4 eq) in water (200 mL) over30 minutes. The reaction mixture was stirred at room temperature for 2hours. The reaction solution was cooled down to 0° C. and was added withsaturated Na₂SO₃ aqueous solution until the reaction color becomescolorless. The stirring was stopped and the organic layer was separatedand the aqueous layer was extracted with EtOAc (3 vol×3). The organiclayers were combined and concentrated in vacuo until the total volumebecomes 3 vol. The resulting solution was extracted with 1N NaOH (3vol×3) and the remaining organic layer was discarded. The combinedaqueous solution was acidified with 6 N HCl until the pH became 1.0. Thesolution was extracted with CH₂Cl₂ (3 vol×5). The combined organic layerwere dried over MgSO₄ and concentrated to afford 8.7 g of the product(compound 4) as a white solid.

¹H NMR (500 MHz, CDCl3) δ 5.84 (s, 1H), 2.23 (t, 2H, J=5.0 Hz),1.51˜1.55 (m, 2H), 0.98 (t, 3H, J=5.0 Hz)

Step 4: Preparation of 3-deutero-(E)-N-cyclopropylhex-2-enamide

To a three-neck 250 mL round bottom flask equipped with mechanicalstirrer and reflux condenser was charged 2-Hexenoic acid-3d (10 g, 0.09mole), IBCF (13 g, 1.1 eq) in CH₂Cl₂ (100 mL, 10 vol). The resultingsolution was cooled down to 0° C. and NMM (13.2 g, 1.5 eq) was addedslowly by controlling the temperature between 0 and 20° C. Then, themixture was allowed to be warmed up to room temperature and stirred for1 hour. To the resulting solution was added cyclopropyl amine (5.9 g,1.2 eq) and the solution was stirred for 2 hours. The reaction mixturewas washed with 1N NaOH (3 vol×2), 1N HCl (3 vol×2), and brine solution(3 vol), and water (3 vol). The organic layer was dried over MgSO₄ andconcentrated to afford the crude product as oil. The crude product wasdissolved with heptane (5 vol) and cooled down to −78° C. with stirring.The precipitated solid was filter and dried to afford 8.7 g of theproduct (compound 5) as a white solid.

¹H NMR (500 MHz, DMSO) δ 7.92 (s, 1H), 5.78 (s, 1H), 2.66˜2.68 (m, 1H),2.08 (t, 2H, J=5.0 Hz), 1.38˜1.42 (m, 2H), 0.87 (t, 3H, J=5.0 Hz), 0.63(t, 2H, J=3.0 Hz), 0.40 (t, 2H, J=3.0 Hz)

Step 5: Preparation of3-deutero-N-cyclopropyl-3-propyloxirane-2-carboxamide

To a three-neck 250 mL round bottom flask equipped with mechanicalstirrer and containing (E)-N-cyclopropylhex-2-enamide-3d (i.e., productfrom Step 4) (10 g, 0.06 mole), urea hydrogen peroxide (25 g, 4 eq), andp-TsOH (12.3 g, 1 eq) in CH₂Cl₂ (100 mL, 10 vol) at 0° C. was addedtrifluoroacetic anhydride (40.9 g, 3 eq) in CH₂Cl₂ (50 mL, 5 vol) over30 minutes. The reaction mixture was heated to 40±5° C. and stirred for3 hours. After cooling to 0° C., the reaction mixture was quenched byadding 6 N NaOH (100 mL, 10 vol) slowly and stirring for 30 minutes. Theorganic layer was separated and washed with brine (5 vol) and water (5vol). The washed organic layer was dried over MgSO₄ and solventevaporated to afford 9.7 g of the epoxide product (i.e., compound 6) aspale yellow oil. The crude product was used for the next step withoutfurther purification.

¹H NMR (500 MHz, DMSO) δ 8.01 (s, 1H), 3.09 (s, 1H), 2.63˜2.65 (m, 1H),1.39˜1.54 (m, 4H), 0.91 (t, 3H, J=5.0 Hz), 0.60 (t, 2H, J=3.0 Hz), 0.45(t, 2H, J=3.0 Hz)

Step 6: Preparation of3-azido-3-deutero-N-cyclopropyl-2-hydroxyhexanamide

To a three necked 250 mL round bottom flask equipped with mechanicalstirrer and reflux condenser containing the epoxide-3d 6 (10 g, 0.06mole) and anhydrous magnesium sulfate (14.1 g, 2.0 eq) in MeOH (100 mL,10 vol) was added sodium azide (15.3 g, 4.0 eq) in one portion. Theresulting mixture was heated to 65±5° C. and stirred for 5 hours. Thereaction mixture was cooled to the room temperature and IPAC (100 mL, 10vol) was added and the mixture was stirred for another 10 minutes. Themixture was filtered through a pad of Celite® to remove insoluble saltsand the resulting clear solution was concentrated to 3 vol. To theresulting solution was added IPAC (170 mL, 17 vol) and the mixture wasstirred for another 10 minutes. Again, the solution was filtered througha pad of Celite® to afford the product, the azide-3d (compound 7), as aclear solution in IPAC (about 200 mL), which was used for the next stepwithout further purification.

¹H NMR (500 MHz, DMSO) δ 7.91 (s, 1H), 6.00 (d, 1H, J=5.0 Hz), 4.03 (d,1H, J=5.0 Hz), 2.66˜2.67 (m, 1H), 1.30˜1.58 (m, 4H), 0.88 (t, 3H, J=5.0Hz), 0.60 (t, 2H, J=3.0 Hz), 0.48 (t, 2H, J=3.0 Hz)

Step 7: Preparation of3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide

To a 500 mL of autoclave hydrogenation reactor equipped with mechanicalstirrer containing the azide-3d 7 (200 mL, 0.05 mole) in IPAC obtainedin the previous step in a hydrogenation reactor was charged Pd/C (10%Pd, water 50%, 0.8 g). The solution was charged with nitrogen (1.0 atm)and released three times and then charged with hydrogen (3.0 atm) andreleased three times. The resulting solution was charged with hydrogen(3 atm) and stirred for 5 hours. After releasing the hydrogen gas, thesolution was purged with nitrogen for 5 minutes. To the resultingsolution was added MeOH(30 ml, 3 vol) and the reaction mixture washeated to 50±5° C. The reaction mixture was filtered through a pad ofcelite to afford a clear solution. The product was isolated byconcentrating the solution at 20±5° C. until 3 vol of the solutionremained. The solid was collected by filtration, washed (IPAC, 3 vol),and dried to give 7.7 g of the title compound (compound 8) as a whitecrystalline solid.

¹H NMR (500 MHz, DMSO) δ 7.70 (s, 1H), 5.31(s, 2H), 3.68 (s, 1H),2.64˜2.66 (m, 1H), 1.10˜1.50 (m, 4H), 0.82 (t, 3H, J=5.0 Hz), 0.59 (t,3H, J=3.0 Hz), 0.45 (t, 3H, J=3.0 Hz)

Step 8: Preparation of(2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide deoxycholate

Deoxycholic acid (15.7 g, 0.75 eq.) was charged to a three-neck 250 mLround bottom flask equipped with mechanical stirrer and containing theracemic (2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide ofstep 7(10 g, 0.05 mole) in THF (100 mL, 10 v). The reaction mixture washeated to 65±5° C. and stirred for 1 hour. The resulting homogeneousmixture was cooled to 23±2° C. over 1 hour, and left at the sametemperature range for 1 hour. The precipitated solids were collected byfiltration, washed with THF (50 mL, 5 vol), and dried to give 12.4 g ofthe salt compound (compound 9) as a white solid. The product has anenatiomeric ratio(ER) of 2:98.

Step 9: Preparation of(2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamidehydrochloride

To a three-neck 250 mL round bottom flask equipped with mechanicalstirrer was charged the dihydrocholate salt (from step 8) and 2-propanol(62 mL, 5 vol). The solution was heated to 75±5° C. and 5 to 6 N HClsolution in IPA (12 mL, 3 eq.) was added slowly with vigorous stirring.The resulting solution was stirred at the same temperature for 1 hourand then cooled down to 23±2° C. The reaction mixture was maintained atthe same temperature for 1 hour. The precipitated solids were collectedby filtration, washed with 2-propanol (36 mL, 3 vol), dried to give 3.0g of the title compound (enantiomeric ratio=0:100) as a white solid. Thedeuterium incorporation was higher than 99% as determined by MS and ¹HNMR.

¹H NMR (500 MHz, DMSO) δ 8.07 (s, 1H), 7.97 (s, 3H), 6.25 (d, 1H, J=5.0Hz), 4.16 (d, 1H, J=5.0 Hz), 2.67˜2.70 (m, 1H), 1.33˜1.46(m, 4H), 0.84(t, 3H, J=5.0 Hz), 0.61 (t, 3H, J=3.0 Hz), 0.53 (t, 3H, J=3.0 Hz).

EXAMPLE 3 Assay for Measuring Epimerization Rate

The deuterated compounds of this invention undergo slow epimerization asfollows:

The epimerization rate was measured according to the following assay.Specifically, 100 μL medium (buffer, rat plasma, dog plasma, or humanplasma) was added into a 96-well deep plate. To the plasma was thenadded 10 μL acetonitrile solution containing a test compound(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)-N—((S)-1-(cyclopropylamino)-1,2-dioxo-3-deutero-hexan-3-yl)octahydrocyclopenta[c]pyrrole-1-carboxamide(at 1 uM or 10 uM) and 1200 μL ethyl acetate into the 96 deep-well plate(2 mL) by using a TomTec liquid handling workstation (Hamden, Conn.,USA). The plate was then covered tightly and shaken with a vortex for 20minutes before it was centrifuged at 3000 rpm for 10 minutes. Aftercentrifuge, 900 μL of the supernatant was transferred to a new V-shape96 deep-well plate using TomTec, and then dried under nitrogen gas (flowrate of 60 L/min) at 25° C. for about 30 minutes. The residue wasreconstituted with 100 μL ethyl acetate, and the solution was againtransferred into the glass inserts in the 96-well plate. 20 uL of thereconstituted solution was injected into LC-MS/MS to determine theamount of the epimers. The LC-MS/MS spectrometer used a ChiralPak ADColumn (4.6×150 mm, 10 μm), a mixture of isopropanol and n-heptane(10:90, 50:50, or 90:10) as the mobile phase, and isopropanol as thewashing solvent. Also used in the MS spectrometer was a deuteratedanalog of the test compound containing 11 deuterium atoms in thecyclohexyl group (C₃₆H₄₂D₁₁N₇O₆, MW 690.47).

The test compound had a mass (M+H, m/z) of 681.36, while itsnon-deuterated analogs (with the same or different chiral configurationsat the deuterated carbon center) had a mass (M+H, m/z) of 680.36. TheirLC-MS/MS spectra showed a fragment of 323.30 (with deuterium) and 322.30(non-deuterated).

At both concentrations (1 uM and 10 uM) and in the same medium (i.e., abuffer, rate plasma, dog plasma, and human plasma), the test deuteratedcompound of formula (I) showed a slower epimerization rate than itsnon-deuterated form buffer, rat plasma, and dog plasma; and a muchslower epimerization rate in human plasma. For instance, in human plasmaand at 1 uM or 10 uM, the deuterated compound epimerized for about 30%in 180 minutes, whereas the non-deuterated form epimerized for almost40%. In addition, in human plasma, the deuterated compound epimerized ata linear rate for 180 minutes, while the non-deuterated form showed anexponential rate of epimerization in the first 60 minutes before itleveled off.

EXAMPLE 4 Assay for Determining IC₅₀ in HCV Replicon Cells

(1S,3aR,6aS)-2-((S)-2-((S)-2-cyclohexyl-2-(pyrazine-2-carboxamido)acetamido)-3,3-dimethylbutanoyl)-N—((S)-1-(cyclopropylamino)-1,2-dioxo-3-deutero-hexan-3-yl)octahydrocyclopenta[c]pyrrole-1-carboxamideand(5S,8S)-3-(5-chloro-2,4-dimethoxyphenyl)-7-((S)-2-(2-cyclohexylacetamido)-3,3-dimethylbutanoyl)-N—((S)-1-(cyclopropylamino)-1,2-dioxo-3-deuterohexan-3-yl)-1-oxa-2,7-diazaspiro[4.4]non-2-ene-8-carboxamidewere used in this assay as described in Lin, C. et al., J. Biol. Chem.,2004, 279: 17508-17514; Lin, K. et al., Antimicrob. Agents Chemother.,2004, 48:4784-4792.

Huh-7 cells harboring an autonomously replicating, subgenomic HCVreplicon of the Con1 strain were maintained in Dulbecco's modifiedEagle's medium (DMEM), 10% heat-inactivated fetal bovine serum (FBS), 2mM L-glutamine, and nonessential amino acids (JRH Biosciences, Lenexa,Kans.), plus 0.25 mg/ml G418 (Invitrogen, Carlsbad, Calif.). Thesubgenomic HCV replicon also encodes a neomycin phosphotransferase,which allows selective growth of HCV replicon-containing Huh-7 cellsover HCV replicon-negative Huh-7 cells in the presence of G418. Theconcentrations of the test compound at which the HCV RNA level in thereplicon cells is reduced by 50% (IC₅₀) or by 90% (IC₉₀) or the cellviability is reduced by 50% (CC₅₀), were determined in HCV Con1subgenomic replicon cells (19) using 4-parameter curve fitting (SoftMaxPro). The replicon cells were incubated with the test compound dilutedin DMEM containing 2% FBS and 0.5% DMSO (without G418) at 37° C. Totalcellular RNA was extracted using an RNeasy-96 kit (QIAGEN, Valencia,Calif.), and the copy number of the HCV RNA was determined in aquantitative, real-time, multiplex reverse transcription-PCR (QRT-PCR,or Taqman) assay. The cytotoxicity of compounds in the HCV repliconcells was measured under the same experimental settings using thetetrazolium-based cell viability assay.

The results show that both test compounds have a Ki of less than 50 nM,and an IC₅₀ (over 5 days) of less than 10.0 uM.

Other Embodiments

It is to be understood that while the invention has been described inconjunction with the detailed description thereof, the foregoingdescription is intended to illustrate and not limit the scope of theinvention, which is defined by the scope of the appended claims. Otheraspects, advantages, and modifications are within the scope of thefollowing claims.

1. A deuterium-enriched α-ketoamido compound of the formula

wherein: D denotes a deuterium atom; R¹ is

in which

is an optionally substituted monocyclic azaheterocyclyl or optionallysubstituted multicyclic azaheterocyclyl, or optionally substitutedmulticyclic azaheterocyclenyl wherein the unsaturatation is in the ringdistal to the ring bearing the R²¹ moiety and to which the—C(O)—N(R²)—CDR³—C(O)—C(O)—NR⁴R⁵ moiety is attached; R²¹ isQ³-W³-Q²-W²-Q¹; wherein Each of W² and W³ is independently a bond, —CO—,—CS—, —C(O)N(Q⁴)-, —CO₂—, —O—, —N(Q⁴)-C(O)—N(Q⁴)-, —N(Q⁴)-C(S)—N(Q⁴)-,—OC(O)NQ⁴-, —S—, —SO—, —SO₂—, —N(Q⁴)-, —N(Q⁴)SO₂—, —N(Q⁴)SO₂N(Q⁴)-, andhydrogen when any of W² and W³ is the terminal group; Each of Q¹, Q²,and Q³ is independently a bond, an optionally substituted aliphatic, anoptionally substituted heteroaliphatic, an optionally substitutedcycloaliphatic, an optionally substituted aryl, an optionallysubstituted heteroaryl, an optionally substituted aralkyl, or anoptionally substituted heteroaralkyl; or hydrogen when any of Q³, Q², orQ¹ is the terminal group, provided that Q² is not a bond when both W³and W² are present; and Each of R², R³, and R⁴, independently, is H or aC₁₋₆ alkyl; and R⁵ is H, alkyl, cycloalkyl, aryl optionally substitutedwith 1-4 alkyl groups, alkylaryl, aryl, amino optionally substitutedwith 1 or 2 alkyl groups.
 2. The compound of claim 1, wherein R²¹ is

in which each of R⁶ and R⁸ is independently a bond; or optionallysubstituted (1,1- or 1,2-)cycloalkylene; or optionally substituted (1,1-or 1,2-)heterocyclylene; or methylene or ethylene, substituted with onesubstituent selected from the group consisting of an optionallysubstituted aliphatic group, an optionally substituted cyclic group andan optionally substituted aromatic group, and wherein the methylene orethylene is further optionally substituted with an aliphatic groupsubstituent; each of R⁷, R⁹, and R¹¹ is independently hydrogen oroptionally substituted aliphatic group; R¹⁰ is an optionally substitutedaliphatic group, optionally substituted cyclic group or optionallysubstituted aromatic group; L is —C(O)—, —OC(O)—, —NR¹¹C(O)—, —S(O)₂—,—NR¹¹S(O)₂—, or a bond; n is 0 or
 1. 3. The compound of claim 2, whereinn is
 1. 4. The compound of claim 2, wherein R⁶ is methylene substitutedwith one substituent selected from the group consisting of an optionallysubstituted aliphatic group, an optionally substituted cyclic group, andan optionally substituted aromatic group.
 5. The compound of claim 4,wherein R⁶ is methylene substituted with isobutyl.
 6. The compound ofclaim 2, wherein R⁷ is hydrogen.
 7. The compound of claim 2, wherein R⁸is methylene substituted with one substituent selected from the groupconsisting of an optionally substituted aliphatic group, an optionallysubstituted cyclic group, and an optionally substituted aromatic group.8. The compound of claim 7, wherein R⁸ is methylene substituted with anoptionally substituted cyclic group.
 9. The compound of claim 8, whereinR⁸ is methylene substituted with cyclohexyl.
 10. The compound of claim2, wherein R⁹ is hydrogen.
 11. The compound of claim 2, wherein L is—CO—.
 12. The compound of claim 2, wherein R¹⁰ is an optionallysubstituted aromatic group.
 13. The compound of claim 12, wherein R¹⁰ isselected from the group consisting of


14. The compound of claim 12, wherein R¹⁰ is optionally substitutedpyrazinyl.
 15. The compound of claim 14, wherein R¹⁰ is 2-pyrazinyl. 16.The compound of claim 2, wherein

is substituted monocyclic azaheterocyclyl.
 17. The compound of claim 16,wherein

is pyrrolidinyl substituted at C-3 position with heteroaryloxy, whereinthe heteroaryl is further optionally substituted with 1-4 halo groups.18. The compound of claim 16, wherein


19. The compound of claim 2, wherein

is optionally substituted multicyclic azaheterocyclyl.
 20. The compoundof claim 19, wherein


21. The compound of claim 20, wherein


22. The compound of claim 2, wherein R² is hydrogen, each of R⁴ and R⁵independently is hydrogen or cyclopropyl.
 23. The compound of claim 2,wherein R³ is propyl.
 24. The compound of claim 2, wherein n is
 0. 25.The compound of claim 2, wherein L is —NR¹¹C(O)— and R¹¹ is hydrogen.26. The compound of claim 2, wherein R¹⁰ is an optionally substitutedaliphatic group.
 27. The compound of claim 26, wherein R¹⁰ is t-butyl.28. The compound of claim 2, wherein the compound is


29. The compound of claim 19, wherein

in which A is —(CHX¹)_(n)—; B is —(CHX²)_(b)—; a is 0 to 3; b is 0 to 3,provided that a+b is 2 or 3; each of X¹ and X² is independently selectedfrom hydrogen, optionally substituted C₁₋₄ aliphatic, and optionallysubstituted aryl; each of Y¹ and Y² is independently hydrogen,optionally substituted aliphatic, optionally substituted aryl, amino, or—OQ⁴; wherein each Q⁴ is independently hydrogen or an optionallysubstituted aliphatic; R²² is an optionally substituted aliphatic, anoptionally substituted heteroaliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, or an optionally substituted heteroaryl.30. The compound of claim 29, wherein R²¹ is optionally substitutedalkylcarbonyl.
 31. The compound of claim 30, wherein R²¹ isaminoalkylcarbonyl, haloalkylcarbonyl, arylalkylcarbonyl,arylalkylcarbonyl, cycloaliphaticalkylcarbonyl, orheterocycloaliphaticalkylcarbonyl, each of which is optionallysubstituted with 1-3 substituents.
 32. The compound of claim 31, whereinR²¹ is heterocycloalkyl-oxycarbonylamino-alkylcarbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,bicycloaryl-sulfonylamino-alkylcarbonyl,aryl-alkoxy-carbonylamino-alkyl-carbonyl,alkyl-carbonylamino-alkyl-carbonyl,aliphatic-oxycarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-aminocarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-carbonylamino-alkyl-carbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,alkyl-aminocarbonylamino-alkyl-carbonyl, orbicycloaryl-aminocarbonylamino-alkyl-carbonyl, each of which isoptionally substituted with 1-3 substituents.
 33. The compound of claim29, wherein R²¹ is

in which each of R⁶ and R⁸ is independently a bond; or optionallysubstituted (1,1- or 1,2-)cycloalkylene; or optionally substituted (1,1-or 1,2-)heterocyclylene; or methylene or ethylene, substituted with onesubstituent selected from the group consisting of an optionallysubstituted aliphatic group, an optionally substituted cyclic group andan optionally substituted aromatic group, and wherein the methylene orethylene is further optionally substituted with an aliphatic groupsubstituent; each of R⁷, R⁹, and R¹¹ is independently hydrogen oroptionally substituted aliphatic group; R¹⁰ is an optionally substitutedaliphatic group, optionally substituted cyclic group or optionallysubstituted aromatic group; L is —C(O)—, —OC(O)—, —NR¹¹C(O)—, —S(O)₂—,—NR¹¹S(O)₂—, or a bond; n is 0 or 1,
 34. The compound of claim 29,wherein R²² is an optionally substituted aliphatic, optionallysubstituted heteroaliphatic, optionally substituted cycloaliphatic,optionally substituted heterocycloaliphatic, optionally substitutedaryl, or optionally substituted heteroaryl.
 35. The compound of claim34, wherein R²² is optionally substituted phenyl, optionally substitutednaphthyl, optionally substituted anthracenyl, optionally substitutednaphthalene, or optionally substituted anthracene.
 36. The compound ofclaim 29, wherein each of X¹, X², Y¹, and Y² is hydrogen, each of a andb is
 1. 37. The compound of claim 36, wherein R²² is an optionallysubstituted aliphatic, optionally substituted heteroaliphatic,optionally substituted cycloaliphatic, optionally substitutedheterocycloaliphatic, optionally substituted aryl, or optionallysubstituted heteroaryl.
 38. The compound of claim 37, wherein R²² is anoptionally substituted phenyl, optionally substituted naphthyl,optionally substituted anthracenyl, optionally substituted naphthalene,or optionally substituted anthracene.
 39. The compound of claim 38,wherein R²¹ is heterocycloalkyl-oxycarbonylamino-alkylcarbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,bicycloaryl-sulfonylamino-alkylcarbonyl,aryl-alkoxy-carbonylamino-alkyl-carbonyl,alkyl-carbonylamino-alkyl-carbonyl,aliphatic-oxycarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-aminocarbonylamino-alkyl-carbonyl,cycloaliphatic-alkyl-carbonylamino-alkyl-carbonyl,heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl,alkyl-aminocarbonylamino-alkyl-carbonyl, orbicycloaryl-aminocarbonylamino-alkyl-carbonyl, each of which isoptionally substituted with 1-3 substituents.
 40. The compound of claim29, wherein


41. The compound of claim 40, wherein the compound is of the structure:


42. The compound of claim 1, wherein the deuterium enrichment is atleast 50% in the compound.
 43. The compound of claim 42, wherein thedeuterium enrichment is at least 80%.
 44. The compound of claim 43,wherein the deuterium enrichment is at least 90%.
 45. The compound ofclaim 44, wherein the deuterium enrichment is at least 99%.
 46. Apharmaceutical composition comprising a pharmaceutically acceptablecarrier and a compound of claim
 1. 47. A method for increasing theconcentration of the active isomer of a pharmaceutical agent in vivo,comprising administering to a patient in need thereof a deuteratedisomer of the pharmaceutical agent in an amount sufficient to confer thepharmaceutical effect.
 48. The method of claim 47, wherein thedeuterated isomer of the pharmaceutical agent is a compound of claim 1.49. A method for enhancing the bioavailability of a compound, comprisingreplacing a hydrogen atom that is bonded to a steric carbon atom in thecompound with a deuterium atom.
 50. The method of claim 49, wherein thedeuterated compound thus obtained is a compound of claim
 1. 51. A methodfor inhibiting HCV protease, comprising contacting HCV protease with acompound of claim
 1. 52. A method for treating a patient suffering fromHCV infection or a condition mediated by HCV protease, comprisingadministering to the patient a pharmaceutically effective amount of acompound of claim
 1. 53. A process for preparing an optically enrichedcompound of Formula 1

wherein: the carbon atoms alpha and beta to the carboxy group arestereocenters; R₁ is independently H, optionally substituted aliphatic,optionally substituted cycloaliphatic, optionally substitutedarylaliphatic, optionally substituted heteroaliphatic or optionallysubstituted heteroarylaliphatic; R′₁ is deuterium such that thedeuterium enrichment is at least 50%; R′₂ is —NHR₂ or —OE; R₂ is H,optionally substituted aliphatic, optionally substituted cycloaliphatic,optionally substituted arylaliphatic, optionally substitutedheteroaliphatic or optionally substituted heteroarylaliphatic; and E isC₁₋₆ alkyl or benzyl; comprising the steps of: a) forming a salt of acompound of Formula 1, and b) crystallizing said salt to give a compoundof greater than 55% enantiomeric excess.
 54. The process of claim 53,wherein R₁ is C₁₋₆ alkyl, and R′₂ is —NHR₂ wherein R₂ is a C₁₋₆ alkyl orC₁₋₆ cycloalkyl.
 55. The process of claim 54, wherein R₁ is propyl andR₂ is cyclopropyl.
 56. The process of claim 53, further comprisingaminating a compound of Formula ii

with an aminating reagent to provide a compound of Formula iii


57. The process of claim 56, wherein the aminating reagent is an azidesalt and the intermediate azido compound is reduced by hydrogenation.58. The process of claim 56, further comprising oxidizing an unsaturatedcompound of Formula i

wherein R′₂ is —NHR₂ or —OE, wherein E is C₁₋₅ alkyl or optionallysubstituted benzyl, with an oxidizing reagent to provide a compound ofFormula ii.


59. The process of claim 58, wherein the oxidizing reagent comprisest-butyl hydroperoxide.
 60. The process of claim 59, wherein theoxidizing reagent further a chiral reagent.
 61. The process of claim 58,wherein the oxidizing reagent is a mixture of samarium (III)isopropoxide, triphenyl arsine oxide, S-(−)1,1′-bi-2-naphthol and 4 Åmolecular sieves.
 62. The process of claim 58, wherein the oxidizingreagent comprises urea-hydrogen peroxide in the presence oftrifluoroacetic anhydride.
 63. The process of claim 62, wherein R′₂ is—OE.
 64. The process of claim 62, wherein R′₂ is —NHR₂.
 65. The processof claim 58, further comprising hydrolyzing the compound of Formula iito give an acid and then converting the acid to an amide compound ofFormula ii wherein R′₂ is —NHR₂.
 66. The process of claim 58, furthercomprising oxidizing a compound of Formula iv

to give the compound of Formula ii.
 67. The process of 66, wherein theoxidation is conducted by using manganese dioxide.
 68. The process of66, further comprising reducing a compound of Formula v

to give the compound of Formula iv.
 69. The process of 68, wherein thecompound is reduced with Red-A1® and then quenched with deuterium oxide.70. A process for preparing a compound of Formula 1

wherein: R₁ is H, optionally substituted aliphatic, optionallysubstituted cycloaliphatic, optionally substituted arylaliphatic,optionally substituted heteroaliphatic or optionally substitutedheteroarylaliphatic; R′₁ is deuterium, R₂ is H, optionally substitutedaliphatic, optionally substituted cycloaliphatic, optionally substitutedarylaliphatic, optionally substituted heteroaliphatic or optionallysubstituted heteroarylaliphatic; and the compound of Formula 1 has anenantiomeric excess of greater than 55%, comprising the steps of: a)oxidation of an unsaturated compound of Formula i

to provide a compound of formula ii

b) reacting a compound of Formula ii with an aminating reagent toprovide a compound of Formula iii

c) forming a salt of a compound of Formula iii with an optically activeorganic acid; d) crystallizing said salt to give a compound of greaterthan 55% enantiomeric excess.
 71. The process of claim 70, wherein thecompound of Formula 1 is(2S,3S)-3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide.
 72. Theprocess of claim 71, wherein the organic acid is L-tartaric acid ordeoxycholic acid.